The overall objective of this project is to elucidate how replication- dependent an replication-independent hi stone synthesis are interrelated during the chromosome replication cycle of proliferating cells and in nonproliferating cells. Such knowledge may be useful in developing procedures with selective toxicity for tumor cells by, as one example, disrupting the balance between histone synthesis and DNA synthesis. The following studies are in progress or have been completed this year. (1) We studied whether the response of the 20 or so members of a histone gen family to DNA replication is the same in different types of replicating cells or whether the response is specific to the cell type. Using four diverse human cell lines we found that the relative expression of seven H2A genes was very similar in all four lines. Thus the evidence suggests that there is a unified histone gene response to replication that is independent of cell type. (2) We studied how a replication-independent histone protein species, H2A.X, results from an apparently replication-dependent gene. We found that part of the answer lies in the regulation of H2A.X gene transcription, which is only slightly modulated by changes in DNA and protein synthesis while the transcription of replication-linked genes is greatly modulated under these conditions. (3) The regulation of H2A.X and H2A.Z gene transcription is being studied by dissection of the essential promoter elements. These two genes both have CCAAT boxes which bind common factors, while the replication-linked H2A.1a gene promote contains similar CCAAT boxes which appear to bind different factors. Thus there appears to be common regulatory elements in the two basal promoters even though the H2A.Z gene contains introns and has polyadenylated transcripts while the H2A.X gen lacks introns and has transcripts that terminate in the conserved histone stemloop. (4) The human H2A.Z gene has been localized to chromosome 4q23 with a pseudogene on chromosome 21. (5)The human H2A.X gene has been localized to chromosome 11q23.